Caspases are activated in a branched protease cascade and control distinct downstream processes in Fas-induced apoptosis.
Two novel synthetic tetrapeptides, VEID-CHO and DMQD-CHO, could selectively inhibit caspase-6 and caspase-3, respectively. We used these inhibitors to dissect the pathway of caspase activation in Fas-stimulated Jurkat cells and identify the roles of each active caspase in apoptotic processes. Affinity labeling techniques revealed a branched protease cascade in which caspase-8 activates caspase-3 and -7, and caspase-3, in turn, activates caspase-6. Both caspase-6 and -3 have major roles in nuclear apoptosis. Caspase-6 cleaves nuclear mitotic apparatus protein (NuMA) and mediates the shrinkage and fragmentation of nuclei. Caspase-3 cleaves NuMA at sites distinct from caspase-6, and mediates DNA fragmentation and chromatin condensation. It is also involved in extranuclear apoptotic events: cleavage of PAK2, formation of apoptotic bodies, and exposure of phosphatidylserine on the cell surface. In contrast, a caspase(s) distinct from caspase-3 or -6 mediates the disruption of mitochondrial membrane potential (permeability transition) and the shrinkage of cytoplasm. These findings demonstrate that caspases are organized in a protease cascade, and that each activated caspase plays a distinct role(s) in the execution of Fas-induced cell death.
Hirata
H
H
Department of Hematology and Oncology, Clinical Sciences for Pathological Organs, Graduate School of Medicine, Kyoto University, Kyoto 606, Japan.
Takahashi
A
A
Kobayashi
S
S
Yonehara
S
S
Sawai
H
H
Okazaki
T
T
Yamamoto
K
K
Sasada
M
M
D034961:Antigens, Nuclear; D017209:Apoptosis; D001324:Autoantigens / Q000378:metabolism; D053148:Caspase 3; D053178:Caspase 6; D053179:Caspase 7; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D018797:Cell Cycle Proteins; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D053938:DNA Fragmentation; D004789:Enzyme Activation; D005434:Flow Cytometry; D006801:Humans; D019169:Jurkat Cells; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins / Q000378:metabolism; D009842:Oligopeptides / Q000494:pharmacology*; D008941:Spindle Apparatus / Q000378:metabolism; D019014:fas Receptor / Q000502:physiology*
D034961:Antigens, Nuclear; D001324:Autoantigens; D018797:Cell Cycle Proteins; D015853:Cysteine Proteinase Inhibitors; C050045:NUMA1 protein, human; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins; D009842:Oligopeptides; C110922:acetyl-aspartyl-methionyl-glutaminyl-aspartyl-aldehyde; C110923:acetyl-valyl-isoleucyl-aspartyl-aldehyde; D019014:fas Receptor; C505407:CASP3 protein, human; C505514:CASP6 protein, human; C505520:CASP7 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053178:Caspase 6; D053179:Caspase 7; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D003546:Cysteine Endopeptidases